Edited by Paul D. Boyer, University of California, Los Angeles, CA, and approved August 15, 2002 (received for review March 13, 2002)

ATP synthase FoF1 (3ß3ab2c10-14) couples an electrochemical proton gradient and a chemical reaction through the rotation of its subunit assembly. In this study, we engineered FoF1 to examine the rotation of the catalytic F1 ß or membrane sector Fo a subunit when the Fo c subunit ring was immobilized; a biotin-tag was introduced onto the ß or a subunit, and a His-tag onto the c subunit ring. Membrane fragments were obtained from Escherichia coli cells carrying the recombinant plasmid for the engineered FoF1 and were immobilized on a glass surface. An actin filament connected to the ß or a subunit rotated counterclockwise on the addition of ATP, and generated essentially the same torque as one connected to the c ring of FoF1 immobilized through a His-tag linked to the or ß subunit. These results established that the c10-14 and 3ß3ab2 complexes are mechanical units of the membrane-embedded enzyme involved in rotational catalysis.

Some have argued that the ATPase may be descended from a pyrophophatase, so this is relevant:

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Proc. Natl. Acad. Sci. USA, 10.1073/pnas.212410399

Pyrophosphate-producing protein dephosphorylation by HPr kinase/phosphorylase: A relic of early life?

In most Gram-positive bacteria, serine-46-phosphorylated HPr (P-Ser-HPr) controls the expression of numerous catabolic genes (10% of their genome) by acting as catabolite corepressor. HPr kinase/phosphorylase (HprK/P), the bifunctional sensor enzyme for catabolite repression, phosphorylates HPr, a phosphocarrier protein of the sugar-transporting phosphoenolpyruvate/glycose phosphotransferase system, in the presence of ATP and fructose-1,6-bisphosphate but dephosphorylates P-Ser-HPr when phosphate prevails over ATP and fructose-1,6-bisphosphate. We demonstrate here that P-Ser-HPr dephosphorylation leads to the formation of HPr and pyrophosphate. HprK/P, which binds phosphate at the same site as the ß phosphate of ATP, probably uses the inorganic phosphate to carry out a nucleophilic attack on the phosphoryl bond in P-Ser-HPr. HprK/P is the first enzyme known to catalyze P-protein dephosphorylation via this phospho-phosphorolysis mechanism. This reaction is reversible, and at elevated pyrophosphate concentrations, HprK/P can use pyrophosphate to phosphorylate HPr. Growth of Bacillus subtilis on glucose increased intracellular pyrophosphate to concentrations (6 mM), which in in vitro tests allowed efficient pyrophosphate-dependent HPr phosphorylation. To effectively dephosphorylate P-Ser-HPr when glucose is exhausted, the pyrophosphate concentration in the cells is lowered to 1 mM. In B. subtilis, this might be achieved by YvoE. This protein exhibits pyrophosphatase activity, and its gene is organized in an operon with hprK.

What we see in nature is that B. subtilis, A. aeolicus, M.tuberculosis, M. genitalium (the smallest genome) , and H. pylor have all 8 parts needed for the F-ATP synthase to function. If this system was truly designed, we would predict that we would find no such evolutionary history for the F-ATP synthase, and we find none. Because of IC, selection would have weeded out any broken ATP synthase, which is why we see it so conserved in all these organisms. Thus an obvious design hypothesis is that the LUCA of all bacteria contained an 8-part ATP synthase which was inteligently designed.

Or a pre-LCA ancestor got by with just a PPase, which is equally successful at generating proton energy gradients but is a heck of a lot simpler than the F1F0 ATPase (and shares some homology to boot).

STRUCTURAL STUDIES OF PROTON TRANSLOCATING PYROPHOSPHATASE Membrane-bound proton translocating pyrophosphatases (H+-PPase) use the energy of pyrophosphate (PPi) hydrolysis to drive proton transport across biological membranes. The formed proton gradient is subsequently used to energize many cellular processes e.g. solute transport and ATP synthesis. The active H+-PPase is a dimer of 60–82 kDa polypeptide monomers, which are predicted to contain 15 transmembrane a-helices. Transmembrane helices are connected by short extracellular turns and longer cytoplasmic loops, three of which are mainly thought to form the active site for PPi hydrolysis. Overall the predicted H+-PPase structure is pretty simple which makes it a good model system for structural and functional elucidation of the mechanism by which pyrophosphate hydrolysis is coupled to proton pumping.

Proteins of the H+-PPase family are found in the vacuolar (tonoplast) membrane of higher plants, algae, and protozoa, and in both bacteria and archaea. They are therefore ancient enzymes. The plant enzymes probably pump one H+ upon hydrolysis of pyrophosphate, thereby generating a proton motive force, postive and acidic in the tonoplast lumen. They establish a pmf of similar magnitude to that generated by the H+-translocating ATPases in the same vacuolar membrane . The bacterial and archaeal proteins may catalyze fully reversible reactions. The enzyme from R.rubrum contributes to the pmf when light intensity is insufficient to generate a pmf sufficient in magnitude to support rapid ATP synthesis.

Eukaryotic members of the H+-PPase family are large proteins of about 770 amino acyl residues with fifteen putative transmembrane a-helical spanners (TMSs). The N-termini are predicted to be in the vacuolar lumen while the C-termini are thought to be in the cytoplasm. These proteins exhibit a region that shows convincing sequence similarity to the regions surrounding the DCCD-sensitive glutamate in the C-terminal regions of the c-subunits of F-type ATPases (TC #3.A.2).

Baltscheffsky is da guy to look at for beginning work on the origin of ATPases; brief online summary:

2. PPi and PPi synthase in the early evolution of biological energy conversion

After our discovery (with von Stedingk, Heldt and Klingenberg) of the first alternative biological electron transport phosphorylation system, leading in bacterial photophosphorylation to PPi rather than to ATP, we have sought evidence for or against the possibility that, in early biological evolution, PPi preceded ATP as the central energy carrier. At present we investigate active site motifs in the proton-pumping PPase family of enzymes (PPase = inorganic pyrophosphatase), to which also bacterial PPi synthase belongs. Of special significance may be certain recurring tetrapeptidyl motifs, which contain 75 - 100 % very early amino acids (Gly, Ala, Asp and Val). These motifs seem to play a central role in PPase function and may be particularly important for obtaining a first detailed picture of the molecular origin and early evolution of biological energy conversion with phosphate compounds. The motifs also show some similarity to corresponding, phosphate binding, regions in both ATP synthases and P-type ATPases.

Note that pyrophosphate can be produced by common inorganic processes.

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So lets review. A de-novo design hypothesis entails:

1. No evolutionary history

2. IC tied in with functional constraint (selection weeding out mutants because of ICness .

Hmm, neither seems quite completely so true for the ATPase, because of the PPase. A simpler, partially sequence-similar system can perform the task. So even for a system older than the flagellum scientists are beginning to get hints indicating that ICness tain't all it's cracked up to be.

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In the near future I want to bring Dembski into the mix. However, unless any relevant criticism of a specific system is brought up, I'm simply going to list them for now. And we can bring up another thread to discuss each system's history. For now, I'm just concerned with listing them.

Well for starters, for the flagellum your reliance on Mike Gene has left you a bit out of date:

MotA and MotB are integral membrane proteins of Escherichia coli that form the stator of the proton-fueled flagellar rotary motor. The motor contains several MotA/MotB complexes, which function independently to conduct protons across the cytoplasmic membrane and couple proton flow to rotation. MotB contains a conserved aspartic acid residue, Asp32, that is critical for rotation. We have proposed that the protons energizing the motor interact with Asp32 of MotB to induce conformational changes in the stator that drive movement of the rotor. To test for conformational changes, we examined the protease susceptibility of MotA in membrane-bound complexes with either wild-type MotB or MotB mutated at residue 32. Small, uncharged replacements of Asp32 in MotB (D32N, D32A, D32G, D32S, or D32C) caused a significant change in the conformation of MotA, as evidenced by a change in the pattern of proteolytic fragments. The conformational change does not require any flagellar proteins besides MotA and MotB, as it was still seen in a strain that expresses no other flagellar genes. It affects a cytoplasmic domain of MotA that contains residues known to interact with the rotor, consistent with a role in the generation of torque. Influences of key residues of MotA on conformation were also examined. Pro173 of MotA, known to be important for rotation, is a significant determinant of conformation: Dominant Pro173 mutations, but not recessive ones, altered the proteolysis pattern of MotA and also prevented the conformational change induced by Asp32 replacements. Arg90 and Glu98, residues of MotA that engage in electrostatic interactions with the rotor, appear not to be strong determinants of conformation of the MotA/MotB complex in membranes. We note sequence similarity between MotA and ExbB, a cytoplasmic-membrane protein that energizes outer-membrane transport in Gram-negative bacteria. ExbB and associated proteins might also employ a mechanism involving proton-driven conformational change.

[...]

The occurrence of significant conformational change in the stator has implications not only for the present-day mechanism but also for the evolution of the flagellar motor. A membrane complex that undergoes proton-driven conformational changes could perform useful work in contexts other than (and simpler than) the flagellar motor, and ancestral forms of the MotA/MotB complex might have arisen independently of any part of the rotor. We queried the sequence database using the sequence of the best-conserved part of MotA (the segment containing membrane segments 3 and 4) from Aquifex aeolicus, a species whose lineage is deeply branched from other bacteria. In addition to the expected MotA homologues, the search returned a protein sequence from the archaeal species Methanobacterium thermoautotrophicum (protein MTH1022) that shows significant sequence similarity not only to MotA but also to the protein ExbB (Figure 9). ExbB is a cytoplasmic-membrane protein that functions in conjunction with ExbD, TonB, and outer-membrane receptors to drive active transport of certain essential nutrients across the outer membrane of Gram-negative bacteria. The energy for this transport comes from the proton gradient across the inner membrane. Thus, MotA and ExbB are both components of systems that tap the proton gradient to do work some distance away (at either the rotor-stator interface or the outer membrane; Figure 9).

Other features also point to a connection between the Mot and Exb systems. MotA functions in a complex with MotB, which as noted contains the critical residue Asp32 near the cytoplasmic end of its single membrane segment. ExbB functions in a complex with ExbD, which likewise has a single membrane segment with a critical Asp residue near its cytoplasmic end (Asp25 in ExbD of E. coli; ref 59). Although ExbB has only three membrane segments in contrast to the four in MotA, the membrane segments that show sequence similarity have the same topology. The protein TonB is also present in the complex with ExbB and ExbD (59, 60) and would provide an additional membrane segment to round out the topological correspondence (Figure 9). ExbB contains a well-conserved Pro residue (Pro141 in E. coli ExbB) that is the counterpart of Pro173 of MotA. Although MotB and ExbD do not share close sequence similarity apart from the critical Asp residue, in certain positions in the membrane segment the residues most common in MotB proteins are also common in ExbD proteins. Finally, like the MotA/MotB complex the ExbB/ExbD complex contains multiple copies of each protein (61). Together, these facts make a reasonable case for an evolutionary connection between the Mot proteins of the flagellar motor and the Exb proteins of outer-membrane transport (and by extension the TolQ/TolR proteins, which are related to ExbB/ExbD but whose functions are less understood).

(bolded)

The number of parts in a flagellum that don't have homologs with different, non-flagellar functions is getting to be rather low; mostly they are filament and shaft proteins, which all may be homologous with each other, and of course nonmotile filaments are known to have a wide degree of uses in bacteria...

So even for systems that are remote from us by 3 billion years there has been some recent progress.

Would any evidence convince evolutionists?The famous British evolutionist (and communist) J.B.S. Haldane claimed in 1949 that evolution could never produce ‘various mechanisms, such as the wheel and magnet, which would be useless till fairly perfect.’10 Therefore such machines in organisms would, in his opinion, prove evolution false. These molecular motors have indeed fulfilled one of Haldane’s criteria. Also, turtles11 and monarch butterflies12 which use magnetic sensors for navigation fulfil Haldane’s other criterion. I wonder whether Haldane would have had a change of heart if he had been alive to see these discoveries. Many evolutionists rule out intelligent design a priori, so the evidence, overwhelming as it is, would probably have no effect.

But none of what you wrote (or linked to) contradicts what I said above. There is no evidence of a pre-PPi synthase, and no evidence that the ATP synthesis molecular machine can be reduced to one component. The synthesis of ATP is a lot more than just inorganic phosphate.

IIRC the ATPase doesn't synthesize the entirety of ATP, it just adds or removes energized phosphates -- Adenosine Triphophsate toe Diphosphate and back.

Rather like what the remarkably simple, 1-component (dimer) PPase does with phosphates:

...and both are coupled to H+ gradients, and use the same basic fold, what a coincidence! It just looks like all of that rotating complexity may be useful and efficiency-increasing add-on rather than an absolutely necessary part of primitive cellular energetics.

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Since the synthesis of diphosphate can be reduced to one subunit, I would say that it is simpler than ATP synthesis, but I wonder if a prediction not unlike the TTSS can be formed, in that it came after or evolved from ATP synthase rather than to it.

You are entitled to your prediction. Regarding TTSS, I am holding out for:

These are predictions that only further data can resolve, however. In general, you'll forgive me for sticking with Baltscheffsky until some IDist (1) acknowledges his existence and (2) explains how they don't greatly weaken the ID argument based on the F1F0 ATPase.

The earliest known H(+)-PPase (proton-pumping inorganic pyrophosphatase), the integrally membrane-bound H(+)-PPi synthase (proton-pumping inorganic pyrophosphate synthase) from Rhodospirillum rubrum, is still the only alternative to H(+)-ATP synthase in biological electron transport phosphorylation. Cloning of several higher plant vacuolar H(+)-PPase genes has led to the recognition that the corresponding proteins form a family of extremely similar proton-pumping enzymes. The bacterial H(+)-PPi synthase and two algal vacuolar H(+)-PPases are homologous with this family, as deduced from their cloned genes. The prokaryotic and algal homologues differ more than the H(+)-PPases from higher plants, facilitating recognition of functionally significant entities. Primary structures of H(+)-PPases are reviewed and compared with H(+)-ATPases and soluble PPases.

Members of the F(o)F(1), A(o)A(1) and V(o)V(1) family of ATP synthases and ATPases have undergone at least two reversals in primary function. The first was from a progenitor proton-pumping ATPase to a proton-driven ATP synthase. The second involved transforming the synthase back into a proton-pumping ATPase. As proposed earlier [FEBS Lett. 259 (1990) 227], these reversals required changes in the H(+)/ATP coupling ratio from an optimal value of about 2 for an ATPase function to about 4 for an ATP synthase function. The doubling of the ratio that occurred at the ATPase-to-Synthase transition was accomplished by duplicating the gene that encodes the nucleotide-binding catalytic subunits followed by loss of function in one of the genes. The halving of the ratio that occurred at the Synthase-to-ATPase transition was achieved by a duplication/fusion of the gene that encodes the proton-binding transporter subunits, followed by a loss of function in one half of the double-sized protein. These events allowed conservation of quaternary structure, while maintaining a sufficient driving force to sustain an adequate phosphorylation potential or electrochemical gradient. Here, we describe intermediate evolutionary steps and a fine-tuning of the H(+)/ATP coupling ratio to optimize synthase function in response to different environments. In addition, we propose a third reversal of function, from an ATPase back to an ATP synthase. In contrast to the first two reversals which required a partial loss in function, the change in coupling ratio required for the third reversal is explained by a gain in function.